In recent times, protein biomarkers have been gaining rapid research interests. Early disease diagnosis and treatment response assessment are made possible with the detection of protein biomarker expression levels in patient samples. Protein biomarkers for cancers, heart diseases and infectious diseases have been reported to date. Characterization of the activity and binding kinetics of the protein biomarkers allow for structural analysis, development of screening assays and lead optimization.
The most commonly used detection methods for biomarker discovery are mass spectroscopy, two-dimensional (2D) western blotting, 2D gel electrophoresis, and enzyme-linked immunosorbent assay (ELISA). In addition, micro-array technology has recently also become an effective alternative for biomarker profiling. Common strategies for antibody microarray detection includes direct target labeling, sandwich assay, and competitive adsorption assay, in which fluorescent labeling, secondary antibodies or enzyme are required in the detection process. Different fluorescently labeled molecules or secondary antibodies are required for each array elements in the antibody array detection. Although mass spectroscopy, 2D western blotting, 2D gel electrophoresis and ELISA detection methods identify whether interactions are present between protein and other molecules, they do not give further information on the binding kinetics of the interactions, which is important to determine drug efficacy. Furthermore, these methods require long detection and preparation processes.
Surface plasmon resonance (SPR) is a sensitive label-free alternative for the detection of biomolecular interactions such as protein-protein interactions. SPR measures the refractive index changes associated with the protein-protein binding occurring at the gold sensing surface. In addition to providing real-time responses for rapid detection, SPR also provides information on protein-protein binding equilibrium and kinetics. Common intensity-based SPR sensors rely on the intensity change produced by the shift of SPR absorption minimum wavelength at resonance. However, such sensors can only provide limited sensor resolution at 10−5 refractive index units (RIUs).